Characteristic expression patterns of allatostatin-like peptide, FMRFamide-related peptide, orcokinin, tachykinin-related peptide, and SIFamide in the olfactory system of crayfish Procambarus clarkii.

The olfactory system plays important roles in various crustacean behaviors. Despite numerous studies on different aspects of the olfactory neural pathway, only the decapod-tachykinin-related peptide (decapod-TRP) has been identified as a neuromodulator in this processing to date. To establish the functions of other related neuropeptides, we initially performed cDNA cloning of FMRFamide-related peptide (FaRP) and allatostatin (AST)-like peptide from the crayfish Procambarus clarkii, followed by in situ hybridization (ISH) analysis of these peptides, along with decapod-TRP, orcokinin, and crustacean-SIFamide. Cloned FaRP cDNA encodes seven copies of C-terminal RN(F/Y)LRFamide-containing peptide, whereas AST-like peptide cDNA comprises 29 copies of AST-like peptide (-YXFGLamide) and three additional putative peptides. ISH analysis of the brain revealed specific expression of crustacean-SIFamide mRNA in most projection neurons (cell cluster 10), and predominant localization of other mRNAs to interneurons. The data suggest that the crustacean-SIFamide neuropeptide is involved in output of the deutocerebrum to the protocerebrum. Double-fluorescence ISH data further disclose that, in cluster 9, orcokinin is coexpressed in decapod-TRP-specific interneurons, whereas AST-like peptide-containing cells do not overlap with orcokinin-expressing cells. On the other hand, FaRP-expressing cells overlap with both orcokinin- and AST-like peptide-specific cells. In cluster 11, where signals for AST-like peptide are absent, a number of interneurons express both decapod-TRP and orcokinin, emphasizing a close relationship between these two factors with regard to olfactory processing, and possibly tactile and/or visual sensory systems. These characteristic expression patterns of neuropeptides support their distinct involvement in the modulation of olfactory processing.

Synthesis, conformational and pharmacological studies of glycosylated chimeric peptides of Met-enkephalin and FMRFa.

Our previous study showed that a chimeric peptide of Met-enkephalin and FMRFamide, YFa (YGGFMKKKFMRFa) not only caused antinociception and potentiated morphine analgesia but also blocked the development of tolerance and physical dependence. In the continuation of that study three chimeric analogues of YFa, [Ser5]YFa, [O-Glu-Ser5]YFa and [O-Gal-Ser5]YFa, were synthesized. To increase the bioavailability and penetration of blood brain barrier (BBB), glycosylated analogues, [O-Glu-Ser5]YFa and [O-Gal-Ser5]YFa, have been synthesized by solid phase peptide synthesis by building block method using anomeric acetate activation method. Circular dichroism studies showed that all the three chimeric peptides are stable and have a propensity for adopting helical conformation in the presence of membrane mimicking solvent. In comparison of parent chimeric peptide YFa, helicity of [Ser5]YFa, [O-Glu-Ser5]YFa and [O-Gal-Ser5]YFa has decreased. Pharmacological studies using tail-flick latency in mice showed that [O-Glu-Ser5]YFa have increased analgesia and bioavailability in comparison of [O-Gal-Ser5]YFa and non-glycosylated analogue [Ser5]YFa. Exhibition of enhanced analgesia by [O-Glu-Ser5]YFa as compared to [O-Gal-Ser5]YFa seems to be due to preference of GLUT-1 transporter system for glucose.

Peptidomic analysis of the larval Drosophila melanogaster central nervous system by two-dimensional capillary liquid chromatography quadrupole time-of-flight mass spectrometry.

Peptides are the largest class of signalling molecules found in animals. Nevertheless, in most proteomic studies peptides are overlooked since they literally fall through the mazes of the net. In analogy with proteomics technology, where all proteins expressed in a cell or tissue are analyzed, the peptidomic approach aims at the simultaneous visualization and identification of the whole peptidome of a cell or tissue, i.e. all expressed peptides with their post-translational modifications. In this paper we describe the analysis of the larval fruit fly central nervous system using two-dimensional capillary liquid chromatography/quadrupole time-of-flight tandem mass spectrometry (LC/Q-TOF-MS/MS. Using the central nervous systems of only 50 larval Drosophila as starting material, we identified 38 peptides in a single analysis, 20 of which were not detected in a previous study that reported on the one-dimensional capillary LC/MS/MS analysis of the same tissue. Among the 38 sequenced peptides, some originate from precursors, such as the tachykinin and the IFamide precursor that were entirely missed in the first study. This clearly demonstrates that the two-dimensional capillary LC approach enhances the coverage of the peptidomic analysis.

Neuropeptide gene families in the nematode Caenorhabditis elegans.

Neuropeptides have diverse roles in the function and development of the nervous system. With the completion of the sequencing of the C. elegans genome, rapid identification of nematode neuropeptide genes is possible. To date, 41 C. elegans neuropeptide genes have been identified. Of these genes, 20 genes, named flp (FMRFamide-like peptide) genes, encode FMRFamide-related proteins (FaRPs). Deletion of one of the flp genes, flp-1, results in several behavioral defects, suggesting that at least one flp gene is not functionally redundant with other flp genes. Twenty-one genes, named neuropeptide-like protein (nlp) genes, encode peptides distinct from the FaRP family. The predicted nlp-1 and nlp-2 neuropeptides have modest similarity to buccalin and myomodulin, respectively. Cellular expression patterns and genetic analysis of flp and nlp genes suggest that neuropeptides in nematodes also have widespread and varied roles in nervous system function.

Comparison of FaRP immunoreactivity in free-living nematodes and in the plant-parasitic nematode Heterodera glycines.

The family of FMRFamide-related peptides (FaRPs) is widely distributed among invertebrates, where the peptides serve as neuromodulators. Published reports indicate that numerous FaRP sequences exist in free-living and animal parasitic nematodes. Using a FMRFamide ELISA, FaRP immunoreactivity was detected in extracts of the soybean cyst nematode, Heterodera glycines, in both sexes and at all developmental stages. HPLC-ELISA results revealed a number of immunoreactive components in H. glycines preparations, and a comparison with extracts of the free-living nematodes Caenorhabditis elegans and Panagrellus redivivus showed significant qualitative differences in FaRP immunoreactivity between the plant parasite and the two free-living nematodes. Total and specific immunoreactivities varied during H. glycines development, with the highest specific activity in juveniles and males, and the highest total activity in mature females. Total female immunoreactivity was located primarily within the mature eggs. A significant portion, however, was associated with the female body, perhaps with egg laying.

Structure, function, and expression of Drosophila melanogaster FMRFamide-related peptides.

In 1977, Price and Greenberg identified the tetrapeptide FMRFamide as a cardioexcitatory molecule from mollusc. Subsequent to this discovery, FMRFamide-related peptides (FaRPs) have been identified in both invertebrates and vertebrates. Peptides in the FaRP family contain a common RFamide C-terminus and act as modulators and messengers of neural and gastrointestinal functions. Like other organisms, Drosophila melanogaster contains several genes that encode for numerous FaRPs. Elucidating the processing and activities of multiple FaRPs encoded in a single precursor is critical to establishing their roles in physiology. In this manuscript, we describe the distribution of FMRFamide immunoreactive materials in the Drosophila central nervous system and gut, and correlate it with the expression of specific FaRPs and their activities. The unique distributions and biological activities of Drosophila FaRPs suggest that the precursors are highly processed and the structurally related peptides are not functionally redundant. The complete distribution of FaRPs in the central nervous system and gut as detected by FMRFamide antisera is not accounted for by the sum of the individual expression patterns of the known Drosophila peptides. Thus, these data suggest that one or more Drosophila FaRPs or structurally related peptides remain to be discovered.

Pharmacology of FMRFamide-related peptides in helminths.

Nervous systems of helminths are highly peptidergic. Species in the phylum Nematoda (roundworms) possess at least 50 FMRFamide-related peptides (FaRPs), with more yet to be identified. To date, few non-FaRP neuropeptides have been identified in these organisms, though evidence suggests that other families are present. FaRPergic systems have important functions in nematode neuromuscular control. In contrast, species in the phylum Platyhelminthes (flatworms) apparently utilize fewer FaRPs than do nematodes; those species examined possess one or two FaRPs. Other neuropeptides, such as neuropeptide F (NPF), play key roles in flatworm physiology. Although progress has been made in the characterization of FaRP pharmacology in helminths, much remains to be learned. Most studies on nematodes have been done with Ascaris suum because of its large size. However, thanks to the Caenorhabditis elegans genome project, we know most about the FaRP complement of this free-living animal. That essentially all C. elegans FaRPs are active on at least one A. suum neuromuscular system argues for conservation of ligand-receptor recognition features among the Nematoda. Structure-activity studies on nematode FaRPs have revealed that structure-activity relationship (SAR) "rules" differ considerably among the FaRPs. Second messenger studies, along with experiments on ionic dependence and anatomical requirements for activity, reveal that FaRPs act through many different mechanisms. Platyhelminth FaRPs are myoexcitatory, and no evidence exists of multiple FaRP receptors in flatworms. Interestingly, there are examples of cross-phylum activity, with some nematode FaRPs being active on flatworm muscle. The extent to which other invertebrate FaRPs show cross-phylum activity remains to be determined. How FaRPergic nerves contribute to the control of behavior in helminths, and are integrated with non-neuropeptidergic systems, also remains to be elucidated.

Actions of nematode FMRFamide-related peptides on the pharyngeal muscle of the parasitic nematode, Ascaris suum.

The endogenous nematode peptides known as FMRFamide-related peptides (FaRPs) and various "classical" transmitters have a range of effects on nematodes that result in changes in behavior, particularly locomotion, including paralysis and inhibition of feeding. This study describes the application of an in vitro pharmacological approach to further delineate the action of a number of FaRP neurotransmitters on feeding behavior. Contraction of Ascaris suum pharyngeal muscle was monitored using a modified pressure transducer system that detects changes in intrapharyngeal pressure and therefore contraction of the radial muscle of the pharynx. The pharynx did not contract spontaneously. However, serotonin (5-HT, 100 microM) stimulated rhythmic contractions and relaxations (pumping) at a frequency of 0.5 Hz. The native nematode peptide, KNEFIRFamide (AF1), inhibited the pumping elicited by 5-HT. The duration of inhibition was concentration-dependent (1-1000 nM) with a threshold of 1 nM (n = 7). KSAYMRFamide (AF8/PF3) also inhibited pharyngeal pumping. There was no observable effect of any of the following nematode peptides on pharyngeal pumping behavior (1-1000 nM; n = 8): AF2, AF3, AF4, AF6, AF16, PF1/CF1, PF2/CF2, or PF4. Thus, interruption of pharyngeal processes, such as feeding, regulation of hydrostatic pressure, and secretion, may provide a new site of anthelmintic action.

Peptidergic control of egg-laying in the cephalopod Sepia officinalis: involvement of FMRFamide and FMRFamide-related peptides.

The peptidergic control of egg-laying was investigated in Sepia officinalis by using a myotropic bioassay. Three myotropic high-performance liquid chromatography fractions were obtained from optic lobe extracts. In the first fraction, FMRFamide (FMRFa) and FLRFa were isolated and sequenced. FMRFa-related peptides then were sought by dotting immunobinding of optic lobes extracts. The four immunoreactive fractions detected revealed the occurrence of FMRFa, FLRFa, FIRFa, and ALSGDAFLRFa predicted by the precursor already cloned from the optic lobes of S. officinalis (J Exp Biol 200:1483-9;1997). These peptides clearly appeared to be involved in the regulation of oocyte transport through the oviduct: the tetrapeptides FMRFa and FLRFa stimulated the contractions, whereas FIRFa and ALSGDAFLRFa lowered the tonus, the frequency, and the amplitude of the contractions. The occurrence of FaRPs in the nervous endings of the accessory sex glands suggested that this peptide family is involved in the regulation of secretory processes of the egg capsule. Indeed, FMRFa modulates the contractions of the main nidamental glands in vitro and, thus, should induce mechanical release of the secretion in vivo during ovulation. These results show that the FaRPs could play an important role in the synchronization of ovulation and egg capsule coating.

FMRFamide-related peptides in the gut of Locusta migratoria L.: a comprehensive map and developmental profile.

The gut tissues and associated nervous system of the African migratory locust, Locusta migratoria, were found to contain FMRFamide-like immunoreactive (FLI) material throughout the five larval instars and 2 weeks into the adult stage in both males and females. FMRFamide-like immunoreactivity associated with the locust gut was described using camera lucida techniques. FMRFamide-like immunoreactivity is observed in the frontal connectives, recurrent nerve, and oesophageal nerves; projections from the ingluvial ganglion onto the anterior midgut, and from the proctodeal nerve onto the hindgut and posterior midgut; in the neuropils of the frontal ganglion, hypocerebral ganglion and ingluvial ganglia; 30 cell bodies in the frontal ganglion; multipolar sensory cells on the foregut; and endocrine-like cells in the gastric caecae and midgut. Radioimmunoassay (RIA) was used to determine the quantities of FLI material in foreguts, gastric caecae, anterior and posterior midguts, and hindgut of first-fifth instar larvae, 1-3- and 14-17-day male and female adult locusts. As expected, as the tissue size (assessed by total protein content) increases, so does the amount of FLI material in each tissue. Normalizing for tissue size reveals significant differences in FLI content among the stages for each tissue tested. Reversed phase-high pressure liquid chromatography (RP-HPLC) followed by RIA has identified four groups of FLI fractions present in the gut, and different members of these groups are present in the various gut tissues.

afp-1: a gene encoding multiple transcripts of a new class of FMRFamide-like neuropeptides in the nematode Ascaris suum.

We have identified a gene, afp-1, that encodes a new subfamily of six FMRFamide-like neuropeptides in the nematode Ascaris suum. The predicted peptides share the C-terminal sequence PGVLRF-NH2 but have different N-terminal extensions. We discuss possible functional roles of these different peptides based upon experiments with Ascaris as well as results from other organisms. Three of the peptides were previously isolated from extracts of A. suum (4) and three other are novel sequences. The translated product of afp-1 is a precursor protein containing two main halves: a C-terminal region containing a series of putative peptides separated by characteristic processing sites and a relatively hydrophobic N-terminal region with no obvious peptides. Although the overall structure of the translated product of afp-1 is similar to flp-1 from C. elegans (18), there is little evidence for homology between the two nematode neuropeptide genes. At least four different transcripts of afp-1 have been identified. These transcripts differ in their 3' and 5' untranslated regions, and one of the transcripts predicts a truncated precursor protein which contains only the C-terminal peptide-containing region.

The effect of FMRFamide analogs on [35S]GTP-gamma-S stimulation in squid optic lobes.

Pharmacological study of Phe-Met-Leu-Phe-amide (FMRFa) receptors is hindered by the lack of selective ligands. The classification of these selective ligands is further hampered by the limited availability of functional assays. In this study, we evaluated several synthetic FMRFa analogs for agonist and antagonist activity by measuring their abilities to produce [35-S]-GTP-gamma-S stimulation or to inhibit FMRFa-induced [35S]-GTP-gamma-S binding in squid optic lobes. Analogs included acetyl-Phe-norLeu-Arg-Phe-amide (acFnLRFa), desamino-Tyr-Phe-Leu-Arg-amide (daYFLRa), desamino Tyr-Phe-norLeu-Arg-Phe-amide (daYFnLRFa), desamino Tyr-Phe-norLeu-Arg-[TIC]-amide (daYFnLR[TIC]a), desamino Tyr-Trp-norLeu-Arg-amide (daYWnLRa), (D)-Tyr-Phe-norLeu-Arg-Phe-amide (D)-YFnLRFa), Phe-Leu-Arg-Phe-amide (FLRFa), and the D-amino acid analogs of FMRFa (D-FMRFa, F-(D)-MRFa and FM-(D)-RFa). For agonist studies, full dose-response curves were generated and analyzed for potency and efficacy (maximal percent effect). FMRFamide as well as analogs ac-FnLRFa, daYFnLRFa, daYFnLR[TIC]a, D-YFnLRFa, FLRFa, and (D)-FMRFa stimulated [35S]-GTP-gamma-S binding. Analogs daYWnLRa, daYFLRa, F-(D)-MRFa, and FM-(D)-RFa failed to stimulate either [35S]-GTP-gamma-S binding or to inhibit FMRFa-induced [35S]-GTP-gamma-S binding. The rank order of potency was daYFnLRFa > or = daYFnLRF[TIC]a > acFnLRFa > (D)YFnLRFa > FLRFa > or = FMRFa >> (D)-FMRFa. The order of efficacy was daYFnLRFa = acFnLRFa = (D)-YFnLRFa > FLRFa = FMRFa > or = (D)-FMRFa > or = daYFnLRF[TIC]a. Peptide analog daYFnLR[TIC]a was less efficacious (59% maximal stimulation) than analogs daYFnLRFa, acFnLRFa, and (D)-YFnLRFa (113-146% maximal stimulation). A maximal concentration of daYFnLR[TIC]a (10 microM) reduced daYFnLRFa, acFnLRFa, and (D)-YFnLRFa induced [35S]-GTP-gamma-S stimulation, indicating that daYFnLR[TIC]a is a partial agonist at the receptor stimulated by the FMRFamide analogs. Analysis of the structural requirements needed for promoting [35S]-GTP-gamma-S binding show that elongation (i.e., daYFnLRFa, D-YFnLRFa) or modification of Phe1 (ac-FnLRFa) leads to increased efficacy and potency. Moreover, elimination of the C-terminal Phe (daYWnLRa, daYFLRa,) leads to a loss of biological activity. However, substitution with L-1,2,3,4 tetrahydroisoquinoline-3-carboxylic acid, a rigid analog of the C-terminal Phe (daYFnLR[TIC]a), leads to decreased efficacy but not loss of potency. The data suggest that immobilization or modification of the C-terminal Phe may produce highly selective and potent FMRFamide antagonists. These results agree with published receptor radioligand studies and indicate that the [35S]GTP-gamma-S assay may be useful in classifying novel FMRFamide-selective ligands.

Contractions associated with the salivary glands of the blood-feeding bug, Rhodnius prolixus: evidence for both a neural and neurohormonal coordination.

The salivary glands of the blood-feeding bug, Rhodnius prolixus, are composed of a single epithelial layer of binucleate cells and a double layer of visceral muscle cells surrounding a large secretory cavity. The saliva contains substances which counteract the hemostasis of the host, and injection of saliva into the host is an essential component of successful and efficient gorging. The muscles surrounding the salivary glands of Rhodnius are under polyneuronal control from the salivary nerve projecting out of the hypocerebral ganglion. The amplitude of contractions induced by neural stimulation is dependent upon both intensity and frequency of nerve stimulation. Serotonin and FMRFamide-related peptides (FaRPs) are delivered in the nerve supply to the salivary glands, and both classes of neuroactive chemicals increase frequency and amplitude of phasic contractions in a dose-dependent manner. A member of the FaRP myosuppressin subfamily, however, inhibits contractions. CRF-related and Leucokinin-like peptides are not delivered in the nerve supply but may be present in the hemolymph during feeding. Leucokinin 1 and Zoone DH (a CRF-related peptide) both induce a dose-dependent increase in basal tonus, with phasic contractions superimposed. Zoone DH is more active than Leucokinin 1. Factors are present in the CNS of Rhodnius which mimic the effects of serotonin and the stimulatory peptides.

Actions of FMRFamide-related peptides on the gCa2+ of the C1 neuron in Helix aspersa.

The endogenous neuropeptides FMRFamide and FLRFamide (tetrapeptides) reversibly reduced a voltage-activated calcium current in the C1 neuron of Helix aspersa by an average of 20%. Two structurally related heptapeptides, pQDPFLRFamide and pQDPFLRIamide, both derived from another precursor protein in this species, did not reduce the current at all.

In vitro release of digestive enzymes by FMRF amide related neuropeptides and analogues in the lepidopteran insect Opisina arenosella (Walk.).

The insect neuropeptides FMRF amide, leucomyosupressin (LMS) and neuropeptide analogues leucosulfakinins (FLSK and LSK II Ser (SO(3)H)), perisulfakinin (PSK), proleucosulfakinin (PLSK), 14A[phi1]WP-I, 542phi1, and 378A[5b]WP-I were assayed for their effects on the release of amylase and protease from the midgut tissue of larvae of Opisina arenosella. In the bioassay, empty midgut tubes ligated at both ends using hair were incubated with insect saline containing neuropeptides/analogues in a bioassay apparatus at 37 degrees C for 30 min. After incubation the contents of the midgut preparations were analyzed for amylase and protease activity. In control experiments, the midgut preparations were incubated in insect saline without neuropeptides. The results of the study reveal that for stimulating amylase release from midgut tissue, the peptides require an FXRF amide (X may be methionine or leucine) sequence at the C-terminal. The presence of HMRF amide at C-terminal of peptides may inhibit the release of amylase. Meanwhile, peptides with both FMRF and HMRF amide sequence at the C-terminal are found to be effective in stimulating protease release. The tetrapeptide segment at the C-terminal probably represent the active core of the neuropeptide.

Action of FMRFamide-like peptides on porcine gastrointestinal motility in vitro.

Mechanical activity was recorded in circular and longitudinal smooth muscle preparations isolated from extensive regions of the porcine gastrointestinal tract in response to the FMRFamide-like neuropeptides F8Famide and A18Famide. In all preparations, the peptides were about equipotent in producing phasic contractions or enhancing spontaneous activity. The most prominent responses were observed in jejunal longitudinal strips which were on the average 91% (+/- 4% SEM, n = 15; 10(-6) M) of the histamine (10(-5) M) responses. The peptide-induced phasic activity was completely abolished by nifedipine but was unaffected by tetrodotoxin, atropine, phentolamine, yohimbine, phenoxybenzamine, propranolol, methysergide, cimetidine, indomethacin, levallorphane or naloxone. Both peptides enhanced acetylcholine-induced contractions. However, bovine ileum and guinea-pig taenia coli was not affected by these peptides. The results indicate that F8F- and A18F-amide contract porcine gastrointestinal smooth muscle by acting directly via non-opioid receptors on L-type calcium channels. In addition an increase of the sensitivity to cholinergic stimulation occurs.

Structure-activity relationships of FMRFamide-related peptides contracting Schistosoma mansoni muscle.

This study reports the potent myoactivity of flatworm FMRFamide-related peptides (FaRPs) on isolated muscle fibers of the human blood fluke, Schistosoma mansoni. The turbellarian peptides YIRFamide (EC50 4 eta M), GYIRFamide (EC50 1 eta M), and RYIRFamide (EC50 7 eta M), all induced muscle contraction more potently than the cestode FaRP GNFFRFamide (EC50 500 eta M). Using a series of synthetic analogs of the flatworm peptides YIRFamide, GYIRFamide and RYIRFamide, the structure-activity relationships of the muscle FaRP receptor were examined. With a few exceptions, each residue in YIRFamide is important in the maintenance of its myoactivity. Alanine scans resulted in peptides that were inactive (Ala1, Ala2, Ala3 and Ala4 YIRFamide; Ala4 and Ala5 RYIRFamide) or had much reduced potencies (Ala1, Ala2 and Ala3 RYIRFamide). Substitution of the N-terminal (Tyr1) residue of YIRFamide with the non-aromatic residues Thr or Arg produced analogs with greatly reduced potency. Replacement of the N-terminal Tyr with aromatic amino acids resulted in myoactive peptides (FIRFamide, EC50 100 eta M; WIRFamide, EC50 0.5 eta M). The activity of YIRFamide analogs which possessed a Leu2, Phe2 or Met2 residue (EC50's 10, 1 and 3 eta M, respectively) instead of Ile2 was not significantly altered, whereas, YVRFamide had a greatly reduced (EC50 200 eta M) activity. Replacement of the Phe4 with a Tyr4 (YIRYamide) also greatly lowered potency. Truncated analogs were either inactive (FRFamide, YRFamide, HRFamide, RFamide, Famide) or had very low potency (IRFamide and MRFamide), with the exception of nLRFamide (EC50 20 eta M). YIRF free acid was inactive. In summary, these data show the general structural requirements of this schistosome muscle FaRP receptor to be similar, but not identical, to those of previously characterized molluscan FaRP receptors.

The modulation of skeletal muscle contraction by FMRFamide-related peptides of the locust.

The external ventral protractor muscle of the VIIth abdominal segment, M234, is a skeletal muscle that possesses receptors that recognize a range of FMRFamide-related peptides and application of these peptides results in an increase in the amplitude of neurally evoked contractions with little or no effect on basal tonus. FLRFamide itself has the same biologic activity as the extended peptides, whereas truncation to LRFamide or RFamide results in a peptide with no biologic activity. The receptors recognizing these extended FLRFamides, which include SchistoFLRFamide, seem to be different from the SchistoFLRFamide receptors found on locust oviduct visceral muscle. SchistoFLRFamide and the non-peptide mimetic, benzethonium chloride (Bztc), increase the frequency and amplitude of miniature endplate potentials, increase the amplitude of neurally evoked excitatory junction potentials, and result in a hyperpolarisation of resting membrane potential. Bztc, however, also abolishes the active membrane response that may explain its ability to decrease neurally evoked contractions.

Evidence for the association of FMRFamide-related peptides with the spermatheca of Locusta migratoria.

The association of FMRFamide-related peptides (FaRPs) with the spermatheca of Locusta migratoria was demonstrated using radioimmunoassay and immunohistochemical techniques. The physiological effects of various FaRPs on the neurally evoked contractions of the spermatheca were also examined. FMRFamide-like immunoreactivity (FLI) was demonstrated in processes and cell bodies situated in the VIIIth (terminal) abdominal ganglion. These included an anterior, central and posterior pair of ventral cell bodies positioned near the midline of the ganglion, in addition to two bilaterally paired dorsal cell bodies in the posterior region of the VIIIth abdominal ganglion. Two axons displaying FLI proceed down the ventral ovipositor nerve (VON) and into the receptaculum seminis nerve which innervates the anterior regions of the spermatheca. FLI was also noted in processes on the spermathecal muscle with the highest density occurring on the spermathecal sac and coil duct. FaRPs applied to the spermathecal muscle included GQERNFLRFamide, NFIRFamide, ADDRNFIRFamide, YGGFMRFamide, FMRFamide, ADVGHVFLRFamide and SchistoFLRFamide (PDVDHVFLRFamide). Dose-dependent physiological effects were only noted for FMRFamide, ADVGHVFLRFamide and SchistoFLRFamide. FMRFamide led to a dose-dependent increase in the amplitude of neurally evoked contractions with a threshold of approximately 5 x 10(-7) M. SchistoFLRFamide, and ADVGHVFLRFamide, had an inhibitory effect, decreasing the amplitude of neurally evoked spermathecal contractions.